Microscopes can receive confusing information if light is bended in an unexpected way; new imaging technology could help fix this problem.
The new approach could prove to be effective in examining subjects that do not scatter light, such as "transparent bodies of zebrafish and the roundworm Caenorhabditis elegans," a Howard Hughes Medical Center news release reported.
"The results are pretty eye-popping," Janelia group leader Eric Betzig said in the news release. "This really takes the application of adaptive optics to microscopy to a completely different level."
"Our technique is really robust, and you don't need anything special to apply our technology. [In the future] it could be a very convenient add-on component to commercially available microscopes," he said.
The researchers turned to adaptive optics techniques used by astronomers and ophthalmologists to correct the light-bending problem. Astronomers achieve this by shining a laser in the same direction of the sky as they plan to observe. The researchers then use a wavefront sensor to see how much the returning light from what is referred to as a "guide star" was distorted.
The researchers created their own "guide star" by "focusing light from the microscope into a glowing point within the sample," the news release reported.
A process called two-photon excitation allowed them to focus the infrared light deep within the transparent tissue to hit their target. The wavefront sensor can then determine how much the light has been distorted.
"In biology, unlike astronomy, the wavefront errors are really complex," Betzig says. "As light from the guide star returns to the sensor, the wavefront gets much bumpier in microscopy than in astronomy. If you fix the guide star at a single point, that bumpiness confuses the sensor, so you don't get a good correction." Furthermore, a correction that works at one point won't be effective at a spot elsewhere in the sample that bends light waves in a different way.
This technique is also used in optics in cases where images of patients' retinas are corrected.
"We combined the descan concept from the ophthalmologists with the laser guide stars of the astronomers, and came up with what amounts to a really good solution for aberrating but non-scattering transparent samples, like the zebrafish," Betzig said.
"We kept on pushing this technology, and it turns out it works," postdoctoral fellow Kai Wang said in the news release. "When we compare the image quality before and after correction, it's very different. The corrected image tells a lot of information that biologists want to know."
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